The present invention, in some embodiments thereof, relates to generation of cytotoxic tumor specific cell lines using Placental Immunoregulatory Ferritin (PLIF) peptides.
Clinical studies on tumor cell-based vaccines are based on the concept that autologous or allogeneic tumor cells express many tumor-associated antigens (TAA). The MHC class I system, along with the endogenous peptides presented on the cell surface are unique markers used by effector CD8+ T cells to discriminate normal cells from diseased cells. MHC class I complexes are constitutively expressed by all nucleated cells in the body. The MHC system includes Ag-processing machinery that processes and presents peptides in the context of MHC molecules on to the cell surface. Thus, cytotoxic T cells made against TAA complexes (TAA/MHC) that mediate anti tumor effects could serve as a novel modality for cancer treatment.
In order to develop immunotherapy for cancer, it is of the utmost importance to have representative target cell lines that present relevant levels of peptides from TAAs on HLA class I molecules. Since HLA-A*0201 is the most common HLA class I molecule in humans, most studies describing the generation of T cells against cancer cell TAAs have focused on HLA-A*0201-restricted peptides.
Several human studies in recent years have demonstrated that the infusion of tumor-specific cytotoxic T cell lines and clones may have a positive clinical effect on diverse malignant diseases, such as colorectal cancer, Hodgkin's lymphoma and nasopharyngeal carcinoma. In most studies documented, the amount of cytotoxic T cell lines required for therapy range from 1×107 to 1×108 per infusion, and most treatment regimens require several cycles of adoptive transfer.
Tumor-specific cytotoxic lymphocytes are usually expanded from peripheral blood mononuclear cells (PBMC) taken from tumor-bearing patients. These are expanded using antigen-presenting cells pulsed with irradiated tumor cells, tumor peptides, tumor lysates or fused tumor cells, resulting in the expansion of MHC class I-restricted cytotoxic T cell lines over several weeks of culture. Tumor-specific cytotoxic T cell lines can also be derived as a subpopulation of tumor-infiltrating lymphocytes by modifying the methodologies, including a purification step based on the selection of CD8 T cells.
In human clinical trials, infusion of tumor-specific T cells derived from tumor-infiltrating lymphocytes or draining lymph nodes has shown limited but encouraging clinical responses in specific settings. Unfortunately, the ability to expand tumor antigen-specific T cells ex vivo from cancer patients is technically difficult due to numerous obstacles, including initiating cultures with low numbers of tumor-specific T cells and the physical inability to obtain tumor-infiltrating lymphocytes from patients with the most common malignancies.
Placenta Immunomodulatory Factor (PLIF) is a protein composed of 165 amino acids. Of these, 117 match the ferritin heavy chain sequence, whereas the C-terminal 48 amino acids (C48) has a sequence which is not related to ferritin. It has been shown that the subcloned recombinant C48 peptide exhibits the bioactivity and therapeutic properties of PLIF [Moroz et al, J. Biol. Chem. 2002, 277, 12901-12905].
PLIF is expressed in the feto-maternal interface in both decidual mononuclear cells and syncytiotrophoblast cells. C48/PLIF binds to macrophages and activated T cells, inducing high levels of IL-10, and acts as a regulatory cytokine. It governs the balance between Th1/Th2 cytokines, which is essential for induction of tolerance during pregnancy. A significantly high correlation was observed between low levels of serum PLIF and the different pathological pregnancy conditions: early pregnancy failures; pregnancies complicated with abortion; intrauterine growth restriction (IUGR); and women at risk for developing pre-eclampsia.
It has been shown that PLIF is upregulated and expressed in malignant cells such as Hodgkin's and non-Hodgkin's lymphoma, acute lymphatic leukemia (ALL), human breast cancer tissues, and breast cancer cell lines (T47D and MCF-7), but not in benign breast disease. Similar to the embryo, PLIF manipulates the cytokine network and immune response, enabling immune escape.
Experiments have been performed to restore T cell immunity and induce rejection of breast cancer by neutralizing C48/PLIF. Rabbit anti-C48 polyclonal antibodies injected intraperitoneally (i.p.) into immune compromised Nude mice engrafted with MCF-7 human breast cancer cells resulted in growth arrest associated with human cell apoptosis and massive intra-tumor lymphocytic infiltration. This was accompanied by activation of INF-γ, thus affecting the cytokine network and leading to breakdown of tolerance.
Synthetic PLIF dimeric peptides are disclosed in U.S. Patent Application No. 61/614,110 for the treatment of cancer.
Additional background art includes U.S. Pat. No. 4,882,270 which discloses a method for detecting breast cancer, by using antibodies against isoferritin placental protein.